摘要： 大豆GmVE2基因是一种NAD（P）H脱氢酶，主要存在于高等植物叶绿体类囊体膜上，可参与氧化应激反应以及叶绿体PSⅠ电子循环传递。为了探究其功能，通过RT-PCR方法从东农50中克隆获得GmVE2基因的CDS全长序列。利用ExPASy-ProtParam等在线软件对GmVE2蛋白序列进行理化性质分析，预测结果表明，GmVE2基因编码区共编码167个氨基酸，分子式为C₉₀₄H₁₃₂₆N₂₀₈O₂₄₆S₁₀，分子质量为19.364 3 ku，总原子数为2 694，理论等电点（pI）为4.78，脂肪指数为87.01；蛋白正负电荷残基数分别为11和23；GmVE2蛋白的不稳定系数和GRAVY值分别为25.02和0.043，说明该蛋白是稳定的疏水性蛋白；蛋白存在跨膜结构、信号肽和磷酸化位点。将线性植物表达载体与目的片段进行连接，成功构建pCambia3300-GmVE2植物表达载体。通过qRT-PCR技术和高效液相色谱（HPLC）分别对动态籽粒及组织基因表达量和维生素E含量进行测定，结果表明，该基因表达量与生育期籽粒维生素E含量呈负相关（-0.951^*），且维生素E含量在绿色组织中显著高于其他组织。本研究对大豆基因GmVE2与维生素E含量的调控关系进行初步研究，为高大豆维生素E含量的大豆新品种选育提供理论和实践基础，为后期研究GmVE2基因功能提供科学依据。

关键词: 大豆, 维生素E, GmVE2基因, 基因克隆, 功能预测, 表达分析

Abstract: Soybean GmVE2 gene is a NAD(P)H dehydrogenase, which is mainly present in thylakoid membranes of chloroplasts of higher plants and can participate in oxidative stress reactions and cyclic PS Ⅰ electron circulation.In order to explore its function, the full-length CDS sequence of GmVE2 gene was cloned from Dongnong 50 by RT-PCR. Use ExPASy-ProtParam and other online software to analyze the physical and chemical properties of the GmVE2 protein sequence.The prediction results showed that the coding region of the GmVE2 gene encoded a total of 167 amino acids, the molecular formula was C₉₀₄H₁₃₂₆N₂₀₈O₂₄₆S₁₀, the molecular weight was 19.364 3 ku, the total number of atoms was 2 694, the theoretical isoelectric point (pI) was 4.78, the fat index was 87.01; the residue bases of positive and negative charges were 11 and 23, respectively. The instability coefficient and GRAVY value of GmVE2 protein were 25.02 and 0.043, respectively, indicating that the protein was a stable hydrophobic protein; the protein had a transmembrane structure, a signal peptide, and a phosphorylation site.The linear plant expression vector was ligated with the target fragment and the pCambia3300-GmVE2 plant expression vector was successfully constructed. The qRT-PCR technology and high performance liquid chromatography (HPLC) were used to determine the gene expression and vitamin E content in dynamic grain and tissue, respectively. The results showed that the gene expression was negatively correlated with the vitamin E content in the growing period grains(-0.951^*), and the vitamin E content was significant higher in green tissues than in other tissues. This study conducted a preliminary study on the regulatory relationship between soybean gene GmVE2 and vitamin E content, providing a theoretical and practical basis for the selection of new soybean varieties with high soybean vitamin E content, and providing a scientific basis for later research on the function of GmVE2 gene.

Key words: Soybean, Vitamin E, GmVE2 gene, Gene cloning, Functional prediction, Expression analysis

中图分类号: 

• S565.03

• Q78